title: Inhibition of energy-producing pathways of HepG2 cells by 3-bromopyruvate.
abstract: 3-BrPA (3-bromopyruvate) is an alkylating agent with anti-tumoral activity on hepatocellular carcinoma. This compound inhibits cellular ATP production owing to its action on glycolysis and oxidative phosphorylation; however, the specific metabolic steps and mechanisms of 3-BrPA action in human hepatocellular carcinomas, particularly its effects on mitochondrial energetics, are poorly understood. In the present study it was found that incubation of HepG2 cells with a low concentration of 3-BrPA for a short period (150 microM for 30 min) significantly affected both glycolysis and mitochondrial respiratory functions. The activity of mitochondrial hexokinase was not inhibited by 150 microM 3-BrPA, but this concentration caused more than 70% inhibition of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 3-phosphoglycerate kinase activities. Additionally, 3-BrPA treatment significantly impaired lactate production by HepG2 cells, even when glucose was withdrawn from the incubation medium. Oxygen consumption of HepG2 cells supported by either pyruvate/malate or succinate was inhibited when cells were pre-incubated with 3-BrPA in glucose-free medium. On the other hand, when cells were pre-incubated in glucose-supplemented medium, oxygen consumption was affected only when succinate was used as the oxidizable substrate. An increase in oligomycin-independent respiration was observed in HepG2 cells treated with 3-BrPA only when incubated in glucose-supplemented medium, indicating that 3-BrPA induces mitochondrial proton leakage as well as blocking the electron transport system. The activity of succinate dehydrogenase was inhibited by 70% by 3-BrPA treatment. These results suggest that the combined action of 3-BrPA on succinate dehydrogenase and on glycolysis, inhibiting steps downstream of the phosphorylation of glucose, play an important role in HepG2 cell death.
authors.affiliation: Laboratório de Bioenergética e Fisiologia Mitocondrial, Programa de Bioquímica e Biofísica Celular, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Av. Carlos Chagas Filho 373-CCS, Bl. D, ss13, 21941-902, RJ, Brazil.
authors.forename: Ana Paula
authors.initials: AP
authors.lastname: Pereira da Silva
authors.forename: Tatiana
authors.initials: T
authors.lastname: El-Bacha
authors.forename: Nattascha
authors.initials: N
authors.lastname: Kyaw
authors.forename: Reinaldo Sousa
authors.initials: RS
authors.lastname: dos Santos
authors.forename: Wagner Seixas
authors.initials: WS
authors.lastname: da-Silva
authors.forename: Fabio C L
authors.initials: FC
authors.lastname: Almeida
authors.forename: Andrea T
authors.initials: AT
authors.lastname: Da Poian
authors.forename: Antonio
authors.initials: A
authors.lastname: Galina
mesh_terms: D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D004734:Energy Metabolism / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism; D006019:Glycolysis / Q000187:drug effects; D006801:Humans; D019344:Lactic Acid / Q000378:metabolism; D008113:Liver Neoplasms / Q000378:metabolism*; D008928:Mitochondria / Q000187:drug effects*; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010101:Oxygen Consumption / Q000187:drug effects; D011773:Pyruvates / Q000494:pharmacology*; D013385:Succinate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D011773:Pyruvates; D019344:Lactic Acid; C017092:bromopyruvate; D000255:Adenosine Triphosphate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D013385:Succinate Dehydrogenase